Breast cancer is a multi-faceted disease that affects a significant number of women (as many as 1 in 8) worldwide. The development of breast tumors is influenced by many factors including genetics, age, and endogenous (ie hormones) as well as exogenous environmental factors. We are interested in determining whether environmental factors interplay with the expression of previously identified cancer susceptibility genes including BRCA1, IGFI, and hTERT to influence the etiology of breast epithelial cell transformation. In studies investigating the function of the breast and ovarian susceptibility gene, BRCA1,we have found that decreased expression of BRCA1 using antisense cDNA resulted in an increase in tumorigenicity and anchorage-independence, as well as a growth advantage in normal growth restrictive conditions, such as low estrogen in cancer cells. In addition, loss of BRCA1 resulted in resistance to apoptotic stimuli induced by hydrogen peroxide and gamma irradiation. To complement this work, we have succeeded in expressing exogenous BRCA1 in cells using a genomic TAR expression vector for use in analysis of the effects of BRCA1 reconstitution in mutant cells. The advantage of this system is that the typical toxicity that is observed with BRCA1 overexpression does not occur. This work will also provide a basis for studying BRCA1 effects in null cells. In related studies, we have defined that in normal breast epithelial cells, BRCA1 can act as a strong transcriptional activator through a specific DNA binding site that we have termed the BRS (Brca1 response sequence). We are currently examining the relevant endogenous promoters that are regulated by BRCA1 and hypothesize that it will play a critical role in transcription-coupled repair mediated by the BRS. Further investigation has revealed that a protein produced from an alternate splicing event may account for most of BRCA1 action in normal cells. In addition we have examined the role of hTERT in mammary cell transformation and have examined the influence of methylation of this promoter in breast cancer cells. Finally, the role of IGFI and the progression of breast carcinoma is being investigated. Preliminary evidence suggests that IGFI may be involved in conversion to the metatstatic phenotype by regulating the expression of critical genes in this process including VEGF.